Isovaleroylbinankadsurin A ameliorates cardiac ischemia/reperfusion injury through activating GR dependent RISK signaling.

Ischemia/reperfusion (I/R) injury is a pathological process caused by reperfusion. The prevention of I/R injury is of great importance as it would enhance the efficacy of myocardial infarction treatment in patients. Isovaleroylbinankadsurin A (ISBA) has been demonstrated to possess multiple bioactivities for treating diseases. However, its protective effect on myocardial I/R injury remains unknown. In this study, the cardiomyocytes hypoxia/reoxygenation (H/R) in vitro model and coronary artery ligation in vivo model were used to examine the protective effect of ISBA. Apoptosis was determined by flow cytometry and Caspase 3 activity. Protein level was determined by Western blot. The mitochondrial viability was examined with mitochondrial viability stain assay. Mitochondrial membrane potential was detected by JC-1 staining and reactive oxygen species (ROS) was stained with 2',7'-dichlorodihydrofluorescein diacetate (DCF-DA). The binding interactions between ISBA and receptors was simulated by molecular docking. Results showed that ISBA effectively protected cardiomyocytes from I/R injury in in vitro and in vivo models. It remarkably blocked the apoptosis induced by H/R injury through the mitochondrial dependent pathway. Activation of the reperfusion injury salvage kinase (RISK) pathway was demonstrated to be essential for ISBA to exert its protective effect on cardiomyocytes. Moreover, molecular docking indicated that ISBA could directly bind to glucocorticoid receptor (GR) and thus induce its activation. Furthermore, the treatment of GR inhibitor RU486 partially counteracted the protective effect of ISBA on cardiomyocytes, consistent with the results of docking.Most attractively, by activating GR dependent RISK pathway, ISBA significantly elevated the cellular anti-oxidative capacity and hence alleviated oxidative damage induced by I/R injury. In conclusion, our study proved that ISBA protected the heart from myocardial I/R injury through activating GR dependent RISK pathway and consequently inhibiting the ROS generation. It provides a valuable reference for ISBA to be developed as a candidate drug for cardiovascular diseases.

An integrated strategy for rapid discovery and identification of quality markers in Guanxin Kangtai preparation using UHPLC-TOF/MS and multivariate statistical analysis.

BACKGROUND: Guanxin Kangtai preparation (GXKT), consisting of Panax ginseng, Panax notoginseng and Ilex pubescens, is a new proprietary Chinese medicines under development for treating coronary heart disease. Like other Chinese medicines, the components of GXKT were complex and the bioactive compounds remained unclear. PURPOSE: To discover bioactive compounds as quality markers (Q-markers) for better quality control of GXKT. STUDY DESIGN: Chinese medicines was separated into fractions. The correlation between chemical information and bioactivity of these fractions were analyzed with multivariate statistical methods to discover bioactive compounds responsible for the actions of Chinese medicine. METHOD: GXKT was separated into fractions by using high-performance liquid chromatography (HPLC). Ultra HPLC coupled with time-of-flight mass spectrometer (UHPLC-TOF/MS) was applied to detect compound information from these fractions to form a chemical database. The bioactivity of these fractions in protecting cardiomyocytes from ischemia/reperfusion injury was examined in H9c2 cells that were exposed to hypoxia followed by reoxygenation (H/R). Then, partial least square model and orthogonal projections to latent structures discriminant analysis were employed to discover bioactive compounds from the chemical database that were positively correlated with the bioactivity of GXKT fractions. Finally, the bioactivity of these compounds was confirmed by bioassay in H9c2 cells. RESULTS: The chemical information of 120 fractions separated from GXKT was detected and extracted by UHPLC-TOF/MS, and a chemical database including 61 high abundance compounds were formed from all fractions. These fractions produced different extent of protective effect to H9c2 cell underwent H/R treatment with cell viability ranging from 33.43% to 74.91%, demonstrating the separation of bioactive compounds among different fractions. The multivariate analysis discovered 16 compounds from GXKT positively correlated with the bioactivity of GXKT. Of these compounds, 6 compounds, i.e.: ginsenoside Rg1, Rb1, Rh1, Rc, ilexsaponin A1, and chikusetsusaponin IVa were chemical identified and also confirmed for their responsibility to the action of GXKT by bioassay. CONCLUSION: Ginsenoside Rg1, Rb1, Rh1, Rc, ilexsaponin A1, and chikusetsusaponin IVa were bioactive compounds and qualified as Q-markers for quality control of GXKT. This research provided a useful reference for the quality research of Chinese medicines.

Six new sesquiterpenoids from Nardostachys chinensis Batal.

Six new sesquiterpenoids, namely nardosinanones J-N and nardoaristolone C, were isolated from the rhizomes and roots of Nardostachys chinensis Batal. Their structures were determined by interpretation of spectroscopic data (HR-ESI-MS, 1D and 2D NMR). A combination of X-ray crystal diffraction, ECD calculation, and Mosher ester methods was employed to determine the absolute configuration of the isolated compounds. Compounds 1-2, 4-6 were evaluated anti-inflammatory activities in LPS-stimulated RAW264.7 macrophages. The results showed that compound 5 obviously inhibited LPS-induced iNOS and COX-2 protein expression compared to single LPS stimulation, which indicated the potential effect to medicate anti-inflammatory.